U6 RNA is an abundant, capped, small nuclear RNA involved in the splicing of pre-messenger RNAs. We recently found that: 1) U6 snRNA is transcribed by RNA polymerase III and this is the first capped snRNA to be synthesized by RNA polymerase III; 2) U6 snRNA is accurately initiated, terminated and capped in vitro; 3) U6 snRNA synthesis is species-specific; 4) Our recent results also showed that at least one factor distinct from RNA polymerase III factors TF-IIIA, IIIB and IIIC, is required for the transcription of a mouse U6 RNA gene. These results indicate the presence of a U6 RNA gene-specific transcription factor; this is analogous to the transcription factor TF-IIIA which is specific to 5S RNA genes. I propose to carry out the following studies: 1) Identify the cis- acting elements of the U6 RNA gene required for the transcription in vitro and in vivo; 2) Identify the factors involved in the transcription of U6 RNA gene; 3) Purify the trans-acting, U6 gene-specific, transcription factor (designated TF-U6) with the U6 RNA gene and identify the binding site(s); and 4) Identify the chemical structure of U6 RNA cap. Since the U6 snRNA is necessary for a fundamental cellular process, and is the first capped RNA synthesized by RNA polymerase III, the studies proposed above are important.